Title : Additional N-glycosylation at
Asn(13) rescues the human
LHbeta-subunit from disulfide-linked aggregation
Abstract :
- CG, LH, FSH , and TSH are a family of heterodimeric glycoprotein hormones that contain a common alpha-subunit , but differ in their hormone-specific beta-subunits
- Despite the considerable homology between LHbeta and CGbeta , we previously demonstrated that, when expressed in GH(3) cells, the secreted form of LHbeta showed mispaired disulfide-linked aggregation in addition to monomer, whereas no aggregation was observed in CGbeta
- To determine the domains which are associated with the LHbeta-aggregation and which prevent CGbeta-aggregation, mutant beta-subunits in glycosylation and carboxy-terminus were expressed in GH(3) cells, and the occurrence of aggregation was assessed by continuous labeling with [35S]methionine/cysteine, immunoprecipitation with anti-h CGbeta serum, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in a non-reducing condition
- No aggregation was seen when N-linked oligosaccharides were attached to the Asn(13) of LHbeta
- Removal of the carbohydrate unit at the Asn(13) of CGbeta caused aggregation, although the amount was less than 10% of monomer
- The carboxy-terminal regions of neither LHbeta nor CGbeta were associated with their aggregation
- Both CGbeta wild-type (WT) and CGbeta lacking N-glycosylation at Asn(13) ( CGbeta- N13 ) showed aggregates in lysate
- However, in contrast to CGbeta- N13 , CGbetaWT revealed no aggregation in medium
- These results indicate that the backbone structure consisting of 114 amino acids and N-linked glycosylation at Asn(30) is involved in the aggregation of LHbeta
- Moreover, N-glycosylation at Asn(13) does not prevent such aggregation, but instead plays an important role in correct folding for both LHbeta- and CGbeta-subunits to be secreted as monomer
Output (sent_index, trigger,
protein,
sugar,
site):
- 0. N-glycosylation, , -, -, Asn(13)
- 1. glycoprotein, , glycoprotein, -, -
- 10. N-glycosylation, , -, -, Asn(13)
- 4. attached, , -, N-linked oligosaccharides, Asn(13)
- 5. Removal, , -, the carbohydrate unit, Asn(13)
- 7. N-glycosylation, , -, -, Asn(13)
- 7. N-glycosylation, , CGbeta, -, Asn(13)
- 7. N-glycosylation, , CGbeta, -, N13
- 9. glycosylation, , -, -, Asn(30)
Output(Part-Of) (sent_index,
protein,
site):
- 4. LHbeta, Asn(13)
- 6. CGbeta, regions
- 6. LHbeta, regions
*Output_Site_Fusion* (sent_index,
protein,
sugar,
site):
- 0. LHbeta, -, Asn(13)
- 10. LHbeta, -, Asn(13)
- 4. LHbeta, N-linked oligosaccharides, Asn(13)
- 5. LHbeta, the carbohydrate unit, Asn(13)
- 7. CGbeta, -, Asn(13)
- 7. CGbeta, -, N13
- 9. LHbeta, -, Asn(30)